Acquired immune deficiency syndrome (AIDS) is a fatal disease. More than 40 million people in the world today are infected with the AIDS virus. The AIDS virus was first identified in 1983, and it has been known by several names and acronyms, including lymphadenopathy-associated virus (LAV), AIDS-related virus (ARV), T-lymphotropic virus III (HTLV-III), and human immunodeficiency virus (HIV). Two distinct families of HIV have been described to date, HIV-1 and HIV-2. The acronym HIV is used herein to refer to human immunodeficiency viruses generically. The AIDS virus is a retrovirus, a virus that uses reverse transcriptase during replication; it has the capacity to replicate within cells of the immune system, causing profound cell destruction.
Current therapeutic regimens have focused on reverse transcriptase and protease inhibitors, but the emergence of mutant drug resistance strains has created a need for more effective and less toxic anti-HIV agents.
Human and related primate immunodeficiency viruses enter a target cell by way of an orchestrated series of recognition events between the invading virus and the target host cell. These include, e.g., interaction of the viral envelope glycoprotein (generically termed “gp120” herein) with the cell surface receptor CD4, followed by subsequent interactions between gp120 and a target cell co-receptor (such as the chemokine receptors, CXCR4, CCR5, etc). The conformational changes brought about by this multi-protein assembly facilitate membrane fusion (e.g., mediated by the “stalk” protein of the viral receptor, the gp41 protein), which is followed by virus entry. Productively infected, virus-producing cells express gp120 at the cell surface; and interaction of gp120 of infected cells with CD4 on uninfected cells results in formation of dysfunctional multicellular syncytia and further spread of viral infection. Thus, interactions between viral envelope proteins, such as gp120, and proteins such as cell surface receptor molecules, e.g., CD4, CXCR4 or CCR5, are particularly attractive targets for interruption of HIV infection and cytopathogenesis, either by prevention of initial virus-to-cell binding or by blockage of cell-to-cell fusion. Virus-free or “soluble” gp120 shed from virus or from infected cells in vivo is also an important therapeutic target, since it may otherwise contribute to noninfectious immunopathogenic processes throughout the body, including the central nervous system.
Several agents have been described that that can block viral envelope (Env)-mediated fusion. One class of such agents is composed of peptides and proteins, including engineered envelope or receptor-derived peptides and proteins. Depending on their size and stability, such inhibitors have the potential to be used as antivirals in vivo or ex vivo. Examples of engineered proteins and peptides that inhibit viral entry include those that target the pre-fusogenic form of gp41 and those that target receptor binding sites on gp120. Examples of nanomolar gp41 inhibitors include the peptides Fuzeon™ (Wild et al. (1994) Proc. Natl. Acad. Sci. U.S.A. 91, 9770-9774), C34 (Chan et al. (1998) Proc. Natl. Acad. Sci. U.S.A. 95, 15613-15617) and N36[Mut]e,g (Bewley et al. (2002) J. Biol. Chem. 277, 14238-14245), and the engineered proteins NCCG-gp41 (Louis et al. (2001) J. Biol. Chem. 276, 29485-29489), N35CCG-N13 (Louis et al. (2003) J. Biol. Chem. 278, 20278-20285) and 5-helix (Root et al. (2001) Science 291, 884-888). Examples of natural proteins that inhibit HIV-1 envelope mediated fusion include cyanovirin-N (CVN) (Boyd et al. (1997) Antimicrob. Agents. Chemother. 41, 1521-1530), scytovirin (Bokesch et al. (2003) Biochemistry 11, 2578-2584), and actinohivin (Chiba et al. (2001) Biochem. Biophys. Res. Commun. 30, 595-601). Interestingly, the three preceding proteins were all isolated from prokaryotic organisms, with CVN and scytovirin coming from the cyanobacteria Nostoc ellipsosporum and Scytonema varium, respectively, and actinohivin from an actinomycete strain.
Microcystis viridis NIES-102 is a freshwater bloom-forming cyanobacterium that was observed to have transient hemagglutinating activity when grown under anaerobic conditions in the laboratory. This activity was traced to a 113 amino acid, 13 kDa protein, termed MVL, the gene for which was cloned and sequenced (Yamaguchi et al. (1999) Biochem. Biophys. Res. Commun 265, 703-708). The present patent describes variants of that protein, and methods for using the protein and the variants, e.g., methods for inhibiting infection by viruses, including HIV.